JP2014506126A - Fat-soluble active ingredient composition containing plant protein-soybean polysaccharide complex - Google Patents

Abstract

  The present invention relates to a) one or more plants selected from the group of proteins suitable for food use a) 0.1 to 70% by weight of one or more fat-soluble active ingredients based on the composition; Protein; and c) for a composition comprising one or more soy soluble polysaccharides, the sum of the protein amount and the polysaccharide amount corresponds to 10-85% by weight, based on the composition in the dry matter. The weight ratio of protein to polysaccharide is selected to be 1: b, where b is included in the range 0.5-15.

Description

Detailed Description of the Invention

  The present invention relates to a composition comprising one or more plant proteins, one or more soy-soluble polysaccharides, and one or more fat-soluble active ingredients. These compositions may be used for enrichment, enhancement and / or coloring of foods, beverages, animal feeds and / or cosmetics. The present invention also refers to the preparation of such compositions. The present invention further refers to a process for producing a beverage by mixing the composition and beverage ingredients. The invention further refers to the beverage obtained by this process.

  Compositions enriching, fortifying or coloring foods, beverages, animal feeds or cosmetics that contain fat-soluble active ingredients such as β-carotene are known in the art. β-carotene is a preferred colored compound because it is very dark in color and is a very nice orange color for the applications described above. Since the final product in which these colorants, nutrients, and / or additives are used is typically an aqueous composition such as a beverage, additional compounds can be added to tolerate the corresponding product. The separation of the fat (oil) phase in the product must be avoided.

  Thus, the fat-soluble active ingredient is often combined with auxiliary compounds such as starch or Aisingrass to prevent phase separation in the final aqueous composition. However, these auxiliary compounds often have an adverse effect on the color and nutritional properties of the final product. Thus, a fat-soluble active ingredient containing improved auxiliary compounds having very good properties with regard to the taste, emulsification, emulsion stability, film-forming ability and / or color of the final product in which it is used It is desirable to develop a new composition.

  Protein has been used as an emulsifier in food products for many years [E. Dickinson, D.D. J. et al. McElements, Molecular basis of protein functionality, in: E.M. Dickinson, D.D. J. et al. McClements (Eds.), Advances in Food Colloids, Black Academic & Professional, London, UK, 1995, pages 26-79]. However, the emulsifying capacity may be lost at or near the isoelectric point, i.e. at a certain protein specific pH at which the net charge and solubility of a particular protein is minimal. In the presence of even higher concentrations of salt, emulsion stability is reduced due to screening for electrostatic repulsion of proteins. Most proteins have an isoelectric point below pH 7. Most foods and beverages are acidic; therefore, the poor stability of emulsions at the isoelectric point limits the applicability of proteins in the food and beverage industry.

  The stability of a protein containing an oil-in-water emulsion depends greatly on the charge density and structure of the emulsifier adsorbed on the emulsion droplet surface. The protein adsorbing layer stabilizes the emulsion film and prevents the combination of droplets. However, protein stabilized emulsions are extremely sensitive to environmental stresses such as pH and ionic strength. [Rungnafar Pongsawamanit, Thepkunya Harnsilawat, David J. et al. McElements, Colloids and Surfaces A: Physicochem. Eng. Aspects, 287, 59-67, 2006]. As the aqueous pH approaches the isoelectric point of the protein and / or the salt concentration is high, the electrostatic repulsion of the protein layer decreases, causing the protein to precipitate, resulting in emulsion droplet coalescence and creaming [Eric Dickinson Soft Matter, 2008, 4, 932-942].

  Proteins as emulsifiers do not function effectively at pH values close to their isoelectric point due to precipitation [N. G. Diftisa, C.I. G. Biriaderisb, V.M. D. Kiossegrou, Food Hydrocolloids 19 (2005) 1025-1031].

  Emulsion stability may be improved by forming protein-polysaccharide conjugates formed through covalent bonds [Eric Dickinson Soft Matter, 2008, 4, 932-942]. Protein-polysaccharide conjugates have improved emulsification and steric stabilization properties, especially under conditions where only the protein has poor solubility [Eric Dickinson Soft Matter, 2008, 4, 932-942].

  Improvements in emulsifying properties of soy protein by conjugation with polysaccharides have been reported [N. Diftis and V. Kiossegrou, Food Chemistry, 81, 1, 2003; Diftis and V. Kiossegrou, Food Hydrocolloids, 20, 787, 2006; G. Difftis, et al. , Food Hydrocolloids, 19, 1025, 2005; Diftis and V. Kiossegrou, Food Chemistry, 96, 228]. Protein-polysaccharide conjugation can improve the emulsification properties of proteins, especially through oil droplet size reduction and emulsion stabilization. These conjugates can be produced by Maillard-type reactions between proteins and polysaccharides, or by other reactions. Xu and Yao (Langmuir 2009, 25 (17), 9714-9720) also describe oil-in-water emulsions prepared from soy protein-dextran conjugates. The conjugate is adsorbed at the interface together with unreacted protein components, increasing the steric stabilization power of the oil droplets. However, Maillard reaction is a time-consuming process and is not suitable for industrial scale reaction. Accordingly, the use of such conjugates remains unsuitable for food and beverage applications.

  Accordingly, there is still a need for compositions comprising fat-soluble active ingredients for enrichment, enhancement and / or coloring of food, beverage, animal feed, cosmetic or pharmaceutical compositions that do not exhibit the above-mentioned problems.

  Thus, for example, a composition of a fat-soluble active ingredient having the desired properties described above, such as very good properties with respect to optical clarity and emulsion stability, and / or improved color strength and color stability (where applicable) It was an object of the present invention to provide a product. It was also an object of the present invention to improve the preparation process of the fat-soluble active ingredient composition.

This purpose is
a) 0.1 to 70% by weight, preferably 0.1 to 30% by weight of one or more fat-soluble active ingredients, based on the composition;
b) one or more plant proteins selected from the group of proteins suitable for food use; and c) one or more soybean soluble polysaccharides,
The total amount of protein and polysaccharide corresponds to 10-85 wt%, preferably 25-85 wt%, more preferably 35-85 wt%, based on the composition of the dry substance,
The weight ratio of protein to polysaccharide was selected to be 1: b, where b was achieved by the composition comprising 0.5-15.

  As used herein, the term “fat-soluble active ingredient” is a vitamin selected from the group consisting of vitamins A, D, E, K, and derivatives thereof; polyunsaturated fatty acids; lipophilic health ingredients; carotenoids And flavorings or odorants and mixtures thereof.

  Suitable polyunsaturated fatty acids (PUFAs) according to the invention preferably have 16 to 24 carbon atoms, in particular 1 to 6 double bonds, preferably 4 or 5 or 6 bis. A mono- or polyunsaturated carboxylic acid having a heavy bond.

  Unsaturated fatty acids can belong to both n-6 and n-3 systems. Preferred examples of n-3 polyunsaturated acids are eicosapenta-5,8,11,14,17-enoic acid and docosahexa-4,7,10,13,16,19-enoic acid; n-6 Preferred examples of polyunsaturated acids are arachidonic acid and γ-linolenic acid.

  Preferred derivatives of polyunsaturated fatty acids are their esters, for example glycerides, in particular triglycerides; particularly preferred are ethyl esters. Particularly preferred are triglycerides of n-3 and n-6 polyunsaturated fatty acids.

  Triglycerides can contain three uniform unsaturated fatty acids or two or three different unsaturated fatty acids. They may also partially contain saturated fatty acids.

  When the derivative is a triglyceride, normally three different n-3 polyunsaturated fatty acids are esterified with glycerin. In a preferred embodiment of the invention, triglycerides are used, 30% of the fatty acid moieties are n-3 fatty acids, 25% of which are long chain polyunsaturated fatty acids. In a further preferred embodiment, commercially available ROPUFA® '30' n-3 Food Oil (DSM Nutritional Products Ltd, Kaiser August, Switzerland) is used.

  In another preferred embodiment of the invention, the PUFA ester is ROPUFA® '75' n-3 EE. ROPUFA '75' n-3 EE is a refined marine oil in the ethyl ester form with a minimum content of n-3 fatty acid ethyl ester of 72%. It is stabilized by mixed tocopherol, ascorbyl palmitate, citric acid and contains rosemary extract.

  In another preferred embodiment of the invention, the PUFA ester is ROPUFA® '10, a purified evening primrose oil comprising at least 9% γ-linolenic acid stabilized by DL-α-tocopherol and ascorbyl palmitate. 'n-6 oil.

  In accordance with the present invention, it may be advantageous to use natural oil (s) containing triglycerides of polyunsaturated fatty acids such as marine oil (fish oil) and / or vegetable oil, but fermented biomass Or oils extracted from genetically modified plants may also be advantageous.

  Preferred oils comprising polyunsaturated fatty acid triglycerides are olive oil, sunflower seed oil, evening primrose seed oil, borage oil, grape seed oil, soybean oil, peanut oil, wheat germ oil, pumpkin seed oil, walnut oil, sesame oil Seed oil, rapeseed oil (canola), cassis seed oil, kiwi seed oil, oil from specific fungi and fish oil.

  Preferable examples of the polyunsaturated fatty acid include linoleic acid, linolenic acid, arachidonic acid, docosahexaenoic acid, eicosapentaenoic acid, and the like.

  Preferred lipophilic health ingredients according to the present invention are selected from resveratrol; Senkyu; coenzyme Q10 (also referred to as “CoQ10”), coenzyme Q9, and / or their reduced form (corresponding ubiquinol) Ubiquinones and / or ubiquinol (one or more components); genistein and / or α-lipoic acid.

  Particularly preferred fat-soluble active ingredients according to the invention are carotenoids, in particular β-carotene, lycopene, lutein, bixin, astaxanthin, apocarotenal, β-apo-8′-carotenal, β-apo-12′-carotenal, canthaxanthin, crypto. Xanthine, citranaxanthin, and zeaxanthin. Most preferred is β-carotene.

  In a preferred embodiment of the invention, the composition is 0.1 to 70% by weight, more preferably 0.1 to 30% by weight, more preferably 0.2 to 20% by weight, based on the total composition. Most desirably, it comprises 0.5 to 15% by weight of one or more fat-soluble active ingredients.

  Preferred plant proteins according to the invention are derived from soybean, lupine (eg L. albus, L. angustifolius or variants thereof), peas and / or potatoes. Protein can be from any part of the plant including fruits (such as soybeans), seeds (including prepared or processed seeds), etc .; or whole wheat flour, or defatted products such as shreds, flakes, etc. May be isolated from the product.

  Particularly preferred for the compositions of the present invention are soy and pea proteins, an even more desirable soy protein is “acid-soluble soy protein” (Soyasour 4000K with a protein content of 60% by weight or more). Most desirable is Soyasour 4000K having a protein content of 80% by weight or more, a water content of 7.5% by weight or less, a fat of 1.5% by weight or less and a pH of 3.6 to 6.4. This is Jilin Fuji Protein Co. Ltd .. Can be supplied from A preferred pea protein source is Cosura SA (Warcoing, Belgium).

  The term “soy soluble polysaccharide” as used herein refers to a soybean soluble polysaccharide having a polysaccharide content of 60% by weight or more. The most desirable soy-soluble polysaccharide has a pH of polysaccharide comprising 70% by weight or more of polysaccharide, 10% by weight or less of protein, 1% by weight or less of fat, 8% by weight or less of moisture, 8% by weight or less of ash, and 3 to 6 Soy soluble polysaccharide. This is Fuji Co. , Ltd., Ltd. Can be supplied from

  It is preferred to select a weight ratio of protein and polysaccharide such as 1: b, provided that b comprises 0.5-15, particularly preferably b is 1-7, more desirably 3-7, Most preferably, it is selected from the range of 4-5.

  In a particularly preferred embodiment of the invention, subsequent heating of the emulsion forms a stable protein-soy soluble polysaccharide emulsifier.

Therefore, the present invention
I) suspending the protein in water;
II) optionally removing undissolved protein from the suspension of step I);
III) mixing one or more soy soluble polysaccharides in a weight ratio of 1: 0.5 to 1:15 protein and polysaccharide;
IV) adjusting the pH to a value comprising 3 to 5 so that a protein-polysaccharide electrostatic complex is formed;
V) adding an organic phase comprising one or more lipophilic active ingredients to the composite;
VI) homogenizing the mixture of step V) by conventional emulsification processes known to those skilled in the art;
VII) heating the emulsion at a temperature comprising 70-95 ° C, preferably 80-90 ° C for at least 45 minutes, preferably at least 1 hour;
VIII) Optionally, drying the emulsion of step VII) also relates to a method of producing a stable emulsifier composition as described above (the method is an amount specified herein) For example).

  Preferred proteins according to the invention are plant proteins as described above.

  The drying step may be carried out by any conventional drying method known to those skilled in the art, preferably spray drying, and / or spray suspension droplets of starch or calcium silicate or silicic acid or calcium carbonate Or a powder trapping process that is captured in an adsorbent bed such as a mixture thereof and subsequently dried.

  The emulsion of step VII) may be used as is or may be dried for later use.

  Homogenization can be performed by standard emulsification techniques such as sonication or high pressure homogenization (800-1200 bar).

  Sonication produces alternating low and high pressure waves in the liquid, resulting in the formation of small vacuum bubbles and severe collapse. This phenomenon, referred to as cavitation, combined with squeezing, acceleration, pressure drop, and impact causes liquid jets that collide at high speeds and strong hydraulic shear forces, resulting in particle collapse throughout the product and Causes dispersion, as well as mixing of reactants. (Encyclopedia of emulsion technology, 1983, Vol 1, P. Walstra, page 57, Ed P. Becher, ISBN: 0-8247-1876-3)

  In the case of a high-pressure homogenization process, the mixture already containing the organic and aqueous phases is passed through a gap in the homogenization valve; this is combined with squeezing, acceleration, pressure drop, and impact, high turbulence and Create shear conditions and cause particle disintegration and dispersion throughout the product. The particle size depends on the operating pressure used in the process and the gap type selected. (Food and Bio Process Engineering, Dairy Technology, 2002, HG Kessler, Ed A. Kessler, ISBN 3-9802378-5-0).

  The most desirable homogenization for practicing the present invention follows (Donsi et al. J. Agric. Food Chem., 2010, 58: 10653-10660), considering the efficiency and high throughput of producing nanoemulsions. High pressure homogenization technology.

  The present invention also relates to a plant protein soy soluble polysaccharide complex obtained by the process as described above, preferably the plant protein is soy protein or pea protein.

  The invention also relates to a composition as described above for enrichment, enhancement and / or coloring of foods, beverages, animal feeds and / or cosmetics, preferably for enrichment, enhancement and / or coloring of beverages. Intended for use.

  Other aspects of the present invention are foods, beverages, animal feeds and cosmetics containing the composition as described above.

  Beverages in which the product form of the invention can be used as a colorant or additive component include, for example, carbonated beverages such as flavored Selzer carbonated water, soft drinks or mineral drinks, and flavored water, fruit juices, fruit punches, and the like It can be a non-carbonated beverage such as a concentrated form of these beverages. They may be natural fruit juice or vegetable juice or an artificial flavor base. Alcoholic beverages and instant beverage powders are also included. In addition, sugar-containing beverages and non-caloric artificial sweetened diet beverages are also included.

  Furthermore, dairy products obtained from natural sources or synthetic are within the scope of food products in which the product forms of the invention can be used as colorants or nutritional ingredients. Typical examples of such products are milk drinks, ice cream, cheese, yogurt and the like. Milk replacement products such as soy milk beverages and tofu products are also included within the scope of this application.

  Also included are confectionery products such as ice creams, jellies, puddings, instant pudding powders, sweet confectioneries such as candy, gums, desserts, etc., containing the product form of the invention as a colorant or additive component.

  Also included are cereals, snacks, cookies, pasta, soups and sauces, mayonnaise, salad dressings and the like containing the product forms of the present invention as colorants or nutritional ingredients. Also included are fruit preparations used for dairy products and cereals.

  The final concentration of one or more fat-soluble active ingredients, preferably carotenoids, especially β-carotene, added to the food product via the composition of the present invention is, for example, about 6 ppm, based on the total weight of the food composition. Etc., preferably 0.1 to 50 ppm, in particular 1 to 30 ppm, more desirably 3 to 20 ppm, depending on the particular food to be colored or fortified and the intended degree of coloring or fortification.

  The food composition of the present invention is preferably obtained by adding a fat-soluble active ingredient in the form of the composition of the present invention to a food product. The compositions of the invention can be used according to methods known per se for the application of water-dispersible solid product forms for the coloring or fortification of foods or pharmaceuticals.

  In general, the composition may be added either as an aqueous stock solution, a dry powder mix, or a pre-blend with other suitable food ingredients, depending on the particular application. Mixing may be performed using, for example, a dry powder blender, a low shear mixer, a high pressure homogenizer, or a high shear mixer, depending on the end use composition. As can be readily appreciated, such expertise is within the technical scope of the expert.

  The present invention provides a beverage comprising the steps of homogenizing the composition according to the present invention and mixing 1 to 50 ppm, preferably 5 ppm of an emulsion with further normal beverage ingredients, based on the fat-soluble content. It also relates to the method of manufacturing.

  The invention further relates to a beverage obtainable by a process for producing a beverage as described above.

Figure 2 shows real-time DLS (dynamic light scattering) results of an emulsion digested over 170 minutes. FIG. 2A shows the ζ-potential of the pea protein solution and supernatant as a function of pH. FIG. 2B shows the ζ-potential of individual pea protein and soy polysaccharide solutions as a function of pH. FIG. 2A shows the ζ-potential of the pea protein solution and supernatant as a function of pH. FIG. 2B shows the ζ-potential of individual pea protein and soy polysaccharide solutions as a function of pH. The visible spectrum of the β-carotene emulsion before and after the addition of FeCl ₃ is shown. The visible spectrum of the β-carotene emulsion before and after the addition of FeCl ₃ is shown. The emulsion was digested with trypsin and pectinase.

  The invention is further illustrated by the following examples, which are not intended to be limiting.

[Example]
[Example 1: Material for producing stable emulsion with soybean protein / soybean soluble polysaccharide]
Soy protein with a protein content of 88% (dry basis) is available from Jilin Fuji Protein Co. Ltd .. (Soyasour 4000K, acid soluble soy protein; ASSP). It had an isoelectric point around pH 4.7. Polysaccharide 70-80% by weight of soybean soluble polysaccharide (SSP) is available from Fuji Co. , Ltd., Ltd. Sourced from.

[Preparation of protein / soybean soluble polysaccharide emulsion]
The preparation of the complex is performed in two steps in situ. First, before homogenization, both products (ASSP and SSP) are mixed and then the emulsion is heated to 80 ° C. to fix the resulting structure. Acid soluble soy protein (ASSP) and soy soluble polysaccharide (SSP) are mixed at pH 3.25 and stirred for 3-4 hours. Soybean oil was then added to the ASSP-SSP aqueous mixture solution. The resulting solution was homogenized with a homogenizer (FJ200-S, Shanghai Specimen Model Co) for 1 minute at room temperature at 10000 rpm and immediately homogenized at 800 bar for 2.5 minutes (AH100D, ATS engineering Inc). Finally, the emulsion was heated at 80 ° C. for 1 hour.

[Particle size measurement]
Each dynamic light scattering (DLS) measurement used an emulsion sample that was freshly diluted at the same pH and NaCl concentration. The measurement was performed on a Malvern Instruments 4700 (Malvern Instruments, Worcs, Malvern PCS7132) and a solid state laser (Compass 315M-100, Coherent Inc .; output about 100 mW, λ = 532 nm). UK). Measurements were performed at 25 ° C. with a fixed scattering angle of 90 °. The measurement time correlation function was analyzed by an Automatic Program equipped with a correlator. The particle size (z-average hydrodynamic diameter, D _h ) was obtained by automatic mode analysis. Two batches of samples were measured and the average data was reported.

[Example 2: Emulsion stability at high salt concentration]
Soy protein ASSP and soy soluble polysaccharide SSP were mixed at pH 3.25. The homogenization conditions are as follows. Protein concentration 5 mg / ml, protein to polysaccharide weight ratio 1: 5, oil volume fraction 10%, homogenization at 800 bar for 2.5 minutes, with or without subsequent heat treatment. After storing overnight at 4 ° C., the pH of the resulting emulsion was adjusted to 5.0 or 6.0 and NaCl was added, then the emulsion was kept at 4 ° C. to investigate long-term stability. The droplet size distribution of the emulsion was measured by dynamic light scattering (DLS). DLS samples were prepared by diluting the emulsion with a freshly prepared aqueous solution having the same pH value and the same salt concentration.

  After the heat treatment, the droplet size distribution of the emulsion does not change significantly compared to the emulsion without heating. However, as shown in Tables 1 and 2, their stability is different. The heated emulsion exhibits long-term stability in pH 5.0 and 6.0 media containing salt.

[Example 3: Stability of emulsion at different pH]
The emulsion has a pH of 3.25, a protein concentration of 5 mg / ml, and a weight ratio of protein to polysaccharide of 1: 5, 10% oil volume fraction, homogenization at 800 bar for 2.5 minutes, heating at 80 ° C. for 1 hour Prepared with or without. The pH of the emulsion was then adjusted to a different value and the emulsion was stored at 4 ° C. for stability. For emulsions that have undergone thermal processing, the emulsion is homogeneous in the pH range 2-8 after storage for 140 and 145 days. However, in the unheated emulsion, creaming appeared in the media at pH 7 and 8 after 20 days of storage; later, creaming also occurred in the unheated emulsion in the media at pH 2 and pH 6. Table 3 also shows an increase in particle size at pH 2 and an increase in pH 5-8; the unheated emulsion increased much more than the heated emulsion before and after storage. The results shown in Table 3 further demonstrate that heat treatment is required to increase the stability of emulsions prepared under high pressure homogenization.

Example 4: Particle size of a heated emulsion freshly prepared with only soy protein or only soy soluble polysaccharide at different pHs
The stability of emulsions prepared with individual acid-soluble soy proteins (ASSP) and individual soy-soluble polysaccharides (SSP) compared to ASSP / SSP complex emulsions was also investigated (see Table 4). . The emulsion was prepared under the conditions of pH 3.25, protein concentration 5 mg / ml or polysaccharide concentration 25 mg / ml, 10% oil volume fraction, homogenization at 800 bar for 2.5 minutes, heating at 80 ° C. for 1 hour. . The emulsion pH was then adjusted to different values and the emulsion was stored at 4 ° C. In the fresh ASSP emulsion, creaming appeared in the pH range 5-8. After 1 week of storage, creaming appeared in all samples except the ASSP emulsion at pH 3.25. This result supports the conclusion that ASSP / SSP composite emulsions are superior to individual ASSP and SSP emulsions.

[Example 5: Effect on homogenization of protein / polysaccharide complex formation pH]
Homogenization was performed at different pH values to investigate the effect of complex formation on the stability of the emulsion. The homogenization conditions are as follows. The ASSP and SSP solutions are adjusted to the desired pH, the ASSP and SSP solutions are mixed at the same pH, protein concentration 5 mg / ml, protein to polysaccharide weight ratio 1: 5, oil volume fraction 10%, at 800 bar. Homogenize for 3-4 minutes. The results shown in Tables 5 and 6 show that homogenization in the pH range 3-4 can produce stable emulsions. In this pH range, ASSP and SSP form an electrostatic complex, where the complex formation is a droplet with an ASSP / SSP complex membrane structure at the oil / water interface and an SSP shell that stabilizes the droplet in aqueous solution. It is suggested that it is essential to generate

[Example 6: Digestion of SSP in emulsion by enzyme]
Emulsion pH 3.25, protein concentration 5 mg / ml, protein to polysaccharide weight ratio 1: 5, oil volume fraction 10%, homogenization at 800 bar for 2.5 minutes, and heating at 80 ° C. for 1 hour Prepared as in Example 1. Next, the soybean polysaccharide in the emulsion was hydrolyzed with pectinase. In order to monitor the particle size change by real-time DLS measurements, hydrolysis was performed at 25 ° C. and pH 5.0 to reduce the hydrolysis rate. During the DLS measurement, 3 μL of 0.5% pectinase solution was added to a polystyrene cuvette containing a diluted emulsion prepared by diluting 5 μL of the original emulsion with 3 mL of pH 5.0 aqueous solution. The droplet size was then measured every 10 minutes for the first 80 minutes and every 15 minutes for another 90 minutes. The data shown (FIG. 1) reveals drop size reduction and stabilization from 262 nm to 219 nm. From the decrease in D _h values before and after digestion, it can be estimated that the polysaccharide layer of the droplet is about 22 nm.

[Example 7: Soybean pea protein solubility in aqueous solution]
Pea protein (Pisane® F9 from Coscro SA Belgium) was dissolved in water at an apparent concentration of 22 mg / mL. The solution was adjusted to pH 1-10 with NaOH or HCl solution. After overnight equilibration, the solution was centrifuged at 5000 rpm or 7800 rpm for 30 minutes. The supernatant was lyophilized and the powder was then weighed to estimate the solubility of pea protein at different pH. The data is shown in Table 7.

  Table 7 shows that the protein concentration is about 2 mg / mL at pH 4 and 5, close to the isoelectric point of pea protein. At pH 1-3 and 7-10, after centrifugation at 5000 rpm, the protein concentration is about 40% higher than after centrifugation at 7800 rpm. This result indicates that the majority of proteins are dispersible aggregates. The solubility of pea protein at pH 3.0 and 4.0 is higher than that of acid-soluble soy protein after centrifugation at 10000 rpm, maintaining the original solution at 20 mg / mL at pH 3.0 and 23 mg / mL at pH 4.0. Much lower.

  A 50 mg / mL pea protein solution was further prepared, and then the solubility was examined by changing the pH to 3.0 and 3.25. The data in Table 8 reveals that about 67% protein aggregates were removed after centrifugation at 5000 rpm for 30 minutes at pH 3.25.

[Example 8: ζ-potential of pea protein solution]
The ζ-potential of the pea protein solution before and after centrifugation at 5000 rpm is not significantly different (FIG. 2A). FIG. 2B shows the ζ-potential of the pea protein and soy polysaccharide solution. The zero ζ-potential of pea protein is about pH 4.8. Pea protein and soy polysaccharide have opposite charges in the pH range of 3.0 to 4.8, in which the pea protein and soy polysaccharide can form an electrostatic complex.

[Example 9: Pea protein / soybean polysaccharide complex emulsion prepared from pea protein solution without centrifugation]
The polysaccharide stock solution at pH 3.25 was diluted with an aqueous solution of the same pH, followed by 0.5 hour stirring. Next, a pH 3.25 pea protein stock solution (no centrifugation) was added. The final protein concentration in the mixed solution was 5 mg / mL, and the weight ratio (WR) of protein to polysaccharide was 1: 5. After stirring the mixed aqueous solution for 3.5 hours, soybean oil was added to make a volume fraction of 10%. The mixture was pre-emulsified using a homogenizer (FJ200-S, Shanghai Specimen Model Co.) for 1 minute at 10000 rpm and immediately emulsified using a high-pressure homogenizer (AH100D, ATS Engineering Inc.) for 4 minutes at 850 bar. Followed by heat treatment at 80 ° C. for 1 hour. After storage at 4 ° C. overnight, the resulting emulsion was adjusted to a different pH number and NaCl was added. Emulsions containing the designed pH number and NaCl concentration were stored at 4 ° C. and examined for stability. The dynamic light scattering (DLS) results of the emulsion are shown in Table 9. After 26 days of storage, the emulsion was homogeneous. After further storage, the emulsion in the medium containing 0.2M NaCl showed creaming. Emulsions in salt-free media at pH 5 and 6 showed a small whey layer at the bottom. The whey layer is less than 10% after 180 days storage compared to the total emulsion volume.

  In order to make the film at the oil-water interface more stable, the heating temperature was changed to 90 ° C. for 1 hour. Other emulsification conditions are the same as described above. After emulsification and heating at pH 3.25, the emulsion was changed to pH 4, 5, 6 and NaCl was added to investigate stability. After 94 days of storage in the pH range 3.25-6, the emulsion was homogeneous in appearance and the droplet size was smaller than 350 nm (Table 10). Creaming appeared in emulsions stored for 94 days in salt media. This result demonstrates that the emulsion is stable in the pH range of 3.25-6 that can be used in salt-free beverages. In the following test, the emulsion was heated at 90 ° C. for 1 hour.

  In addition to emulsification at pH 3.25, emulsification is also at pH 3.0, 3.5, 3.75, and 4.0. But I did it. The resulting emulsion produced in the pH range 3.5-4.0 was unstable due to reduced pea protein solubility in this pH range. Emulsions produced at pH 3.0 are not as stable as pH 3.25.

[Example 10: Pea protein / soybean polysaccharide complex emulsion prepared from a pea protein solution centrifuged at pH 3.25]
Next, in order to produce a stable emulsion in a salt medium, the emulsion is produced from a protein solution that has been centrifuged at pH 3.25 to remove aggregates that cannot be decomposed. The final protein concentration in the aqueous solution was approximately 4 mg / mL and the polysaccharide was 25 mg / mL. Other conditions are the same as above. The resulting emulsion droplet size is shown in Table 11. The emulsion is homogeneous after 87 days storage at pH 5 and 6 with or without 0.2 M NaCl; the droplet size (D _h ) does not change significantly after storage in different media.

Example 11 Pea Protein / Soybean Polysaccharide Complex Emulsion Prepared from Pea Protein Solution Centrifugated at pH 6.8
The centrifugation pH was changed to increase the utilization of pea protein and to obtain a stable emulsion in salt medium. The original pea protein solution has a pH of 6.8 and the original soy polysaccharide solution has a pH of 5.3. The protein solution is centrifuged at pH 6.8, at which the protein utilization is about 73%. The protein solutions were then mixed with the soy polysaccharide solution at pH 5.3 (without pH adjustment) or they were mixed at pH 7.0 (with pH adjustment). The protein / polysaccharide mixture was further adjusted to pH 3.75. Table 12 shows that there is no precipitation at pH 3.75, suggesting that the polysaccharide may protect the protein from precipitation. Mixtures mixed at pH 7.0 and then changed to pH 3.75 have smaller particle size and greater strength, suggesting better complexation between the protein and polysaccharide. When mixed at pH 5.3, protein aggregates can inhibit protein binding to polysaccharides. The mixture of Table 12 is used to produce an emulsion at pH 3.25; the droplet size is shown in Table 13. Since the pea protein / soybean polysaccharide complex solution mixed at pH 7.0 produced smaller droplets, the following conditions were employed to produce an emulsion in the following test. The pea protein solution at pH 6.8 is centrifuged at 5000 rpm for 30 minutes and the resulting supernatant pH and soy polysaccharide solution pH are adjusted to pH 7.0, respectively, and the protein and polysaccharide at pH 7.0 are adjusted. Solution mixing followed, and then the mixture was changed to emulsified pH.

  Different weight ratio (WR) pea protein and soy polysaccharide complex solutions were further investigated. The data in Table 14 further support that complexation can break up aggregates of individual proteins and individual polysaccharides to form smaller complex particles.

[Example 12] Pea protein / soybean polysaccharide complex emulsion prepared from a pea protein solution centrifuged at pH 6.8. Both pea protein and soy polysaccharide solution were adjusted to pH 7.0 before mixing.

  In the following test, the procedure for producing the emulsion is as follows. The aqueous solution of pea protein (pH 6.8) was centrifuged at 5000 rpm for 30 minutes. The pea protein and soy polysaccharide solutions were each adjusted to pH 7.0, then mixed and stirred for 2 hours. The mixture was further adjusted to the emulsifying pH. After stirring for another 4 hours, soybean oil was added to the 10% volume fraction. The mixture was pre-emulsified using a homogenizer at 10,000 rpm for 1 minute, immediately emulsified using a high-pressure homogenizer at 800 bar for 3 minutes, followed by heat treatment at 90 ° C. for 1 hour. After storage overnight at 4 ° C., the resulting emulsion was adjusted to a different pH number and NaCl was added. Emulsions containing the designed pH number and NaCl concentration were stored at 4 ° C. and examined for stability.

[(1) Effect of emulsification pH]
Complex emulsions were produced in the pH range 2.5-7. The DLS results (Table 15) show that the emulsions produced in the pH range 3.5-4.25 are stable in 0.2M NaCl addition medium at pH 5-6; the emulsion is homogeneous after storage there were. In the pH range 3.5-4.25, as shown in FIG. 2, the electrostatic attraction between proteins and polysaccharides is stronger, which favors complex formation. In the following test, pH 3.75 was used as the emulsification pH.

[(2) Effect of weight ratio (WR) between pea protein and soybean polysaccharide]
The protein concentration was fixed at 5 mg / mL and the polysaccharide concentration was changed from 2.5 to 30 mg / mL, ie the WR was changed from 2: 1 to 1: 6. Table 16 shows that when WR is in the range of 1: 2 to 1: 6, the droplet size is not significantly affected by the environment, and the oil droplets are coated with sufficient polysaccharide. It is suggested. WR 1: 4 and 1: 5 were selected for further testing.

[(3) Effect of high-pressure homogenization (HPH)]
The homogenization pressure was changed from 800 to 1200 bar. The data in Table 17 demonstrates that pressure has no significant effect on emulsion droplet size and stability. In the following tests, HPH conditions were fixed at 800 bar for 3 minutes.

[(4) Effect of heat treatment]
An emulsion produced from a WR 1: 4 complex with a protein concentration of 5 mg / mL at pH 3.75 and 800 bar for 3 minutes was divided into two parts. One was heated at 90 ° C. for 1 hour and the other was not heated. The emulsion was then changed from pH 2 to 8 and 0.2 M NaCl was added. The data in Table 18 suggests that the heated emulsion is stable to changes in pH and salt concentration. Heat induces protein denaturation and can form an irreversible oil-water interface film composed of pea protein and soy polysaccharides. During storage in different media, the non-heated emulsion showed creaming in all media containing salt and creamed in media with pH 7 and 8 without salt. In contrast, heated emulsions are homogeneous in all media with or without salt. Table 18 also shows that the unheated emulsion is stable in the pH range 3-5, the emulsion is homogeneous after storage, and the droplet size does not change. This result suggests that unheated emulsions can encapsulate heat sensitive lipophilic bioactive compounds and the emulsions can be used in salt-free beverages.

[Example 13] A pea protein / soybean polysaccharide complex emulsion prepared from a pea protein solution without centrifugation. Both pea protein and soy polysaccharide solution were adjusted to pH 7.0 before mixing.

[1] Emulsion prepared from pea protein solution without centrifugation. ]
The pea protein and soy polysaccharide solutions were each adjusted to pH 7.0, then mixed and stirred for 2 hours. The protein concentration was 5 mg / mL and the WR was 1: 4. The mixture was further adjusted to the emulsifying pH. After stirring for another 4 hours, soybean oil was added to the 10% volume fraction. The mixture was pre-emulsified using a homogenizer at 10000 rpm for 1 minute, immediately emulsified using a high-pressure homogenizer at 800 bar for 4 minutes, followed by heat treatment at 90 ° C. for 1 hour. After storage overnight at 4 ° C., the resulting emulsion was adjusted to a different pH number and NaCl was added. Emulsions containing the designed pH number and NaCl concentration were stored at 4 ° C. and examined for stability (Table 19). The stability of emulsions produced at pH 3.5 and 3.75 in different media was further investigated (Table 20). The data in Tables 19 and 20 show that emulsions prepared from non-centrifuged pea protein solutions are also stable to changes in pH and salt concentration.

[2] Emulsions prepared with 20% and 30% oil volume fractions. ]
The pea protein non-centrifugation solution and soy polysaccharide solution were each adjusted to pH 7.0, then mixed and stirred for 2 hours. The protein concentration was 5 mg / mL and the WR was 1: 4. The mixture was further adjusted to pH 3.5 and then soybean oil was added to the 20% and 30% volume fractions. The mixture was emulsified at 800 bar for 4 minutes, followed by heat treatment at 90 ° C. for 1 hour. The data in Table 21 suggests that the droplet size increases with oil content and that emulsion stability decreases with increasing oil content.

Example 14: Emulsifying capacity and stability of individual pea proteins
Individual pea protein solutions at a concentration of 5 mg / mL were adjusted to pH 3, 4, 5, 6, and 7, followed by emulsification. Emulsions produced at pH 4, 5, and 6 showed creaming immediately after emulsification. The emulsions produced at pH 3 and 7 have small droplet sizes of 290 and 358 nm before heating and 273 and 398 nm, respectively, after heating. The heated and unheated emulsions produced at pH 3 and 7 were changed from pH 2 to 8 and NaCl was added. DLS results show a rapid increase in droplet size for freshly prepared samples (data not shown); all samples except emulsions produced at pH 3.0 have a creaming of 1 without pH change and addition of salt It showed that it appeared within a week.

  The above results support the conclusion that pea protein / soy polysaccharide complex emulsions are superior to individual pea protein and individual soy polysaccharide emulsions.

[Example 15: Encapsulation of β-carotene in complex emulsion droplets]
β-carotene was added to soybean oil at a concentration of 1 mg / mL. The oil phase was heated at 70 ° C. for 2 hours under a nitrogen atmosphere to dissolve β-carotene. The emulsion was prepared at pH 3.75, 800 bar for 4 minutes with 10% oil volume fraction, 5 mg / mL pea protein, and 20 mg / mL polysaccharide aqueous phase. After emulsification, β-carotene was encapsulated in droplets; β-carotene was 0.1 mg / mL in the emulsion. The resulting emulsion was adjusted to a different pH and examined for stability. All emulsions were homogeneous in appearance after 11 days of storage at different pHs, but the DLS results shown in Table 22 show that capsules in pH ranges 3, 4, and 5 where the droplet size does not change significantly. This suggests that the stabilized emulsion is stable.

To further demonstrate the stability of encapsulated β-carotene, the visible spectrum of the β-carotene emulsion was measured before and after the addition of FeCl ₃ (see FIG. 3). For this experiment, the encapsulated material was incubated with FeCl ₃ . An emulsion was prepared by mixing 5 μL of β-carotene emulsion and 3 ml of water and adding 20 μL of FeCl ₃ at a concentration of 10 mg / ml to it. The final β-carotene concentration was 1.67 × 10 ⁻³ mg / mL. The blank solution for measurement contains the same components at the same concentration but does not contain β-carotene. This experiment shows that the absorption band characteristic of β-carotene emulsion does not change after the addition of FeCl ₃ , demonstrating that the charged β-carotene cannot react with FeCl ₃ , so the emulsion is preserved It is confirmed that β-carotene can be protected from oxidation.

To further determine the activity of the β-carotene emulsion, pectinase and trypsin were used, respectively, to release β-carotene by digestion of soybean polysaccharide and pea protein. The process is as follows. 5 μL of β-carotene emulsion was added to 2.6 mL of pH 8.2 Tris buffer (20 mM), and 400 μL of trypsin solution prepared by dissolving trypsin in Tris buffer at a trypsin concentration of 2 mg / mL was added. The β-carotene concentration in the mixture was 1.67 × 10 ⁻³ mg / mL. After measuring the visible spectrum, 20 μL of 10 mg / mL FeCl ₃ was added to the mixture. These results (FIG. 4) indicate that the absorption band characteristic of β-carotene does not change in the presence of FeCl ₃ . The mixture was then adjusted to pH 5.0 and 40 μL pectinase solution and 20 μL 10 mg / mL FeCl ₃ were added to the mixture. The visible spectrum then shows that the absorption band characteristic of β-carotene has disappeared, demonstrating that the released β-carotene can react with FeCl ₃ . During spectral measurements, the blank solution contains the same components at the same concentration but does not contain β-carotene. Control experiments confirmed that the characteristic absorption band of β-carotene did not change in the presence of FeCl ₃ in the β-carotene emulsion when only trypsin or pectinase was added.

[Example 16: Encapsulation of vitamin E in complex emulsion droplets]
Initially, pure vitamin E was used as the oil phase to produce an emulsion. The emulsion was prepared at pH 3.25, 800 bar for 3 minutes with 10% oil volume fraction, 5 mg / mL pea protein, and 20 mg / mL polysaccharide aqueous phase. The emulsion was heated at 90 ° C. for 1 hour. The DLS results shown in Table 23 indicate that the emulsion is not stable in salt media.

  Considering the viscosity of vitamin E, the oil phase was changed to a mixture of 40% vitamin E and 60% soybean oil. Other conditions are the same as above. An emulsion was produced at pH 3.75. The data in Table 24 indicates that the droplet size is not sensitive to changes in pH and salt concentration, implying that the emulsion is stable in these media.

[Example 17: Effect of weight ratio (WR) of soybean protein and soybean polysaccharide on stability of soybean protein / soybean polysaccharide emulsion]
Table 25 shows soy protein / soy polysaccharide emulsions prepared from different weight ratios (WR) of soy protein and polysaccharides. The data in Table 24 reveals that emulsions prepared from WR 1: 2 to 1: 6 complexes are stable to changes in pH and salt concentration, and long-term storage.

[Example 18] Effect of weight ratio (WR) of pea protein and soybean polysaccharide on stability of pea protein / soybean polysaccharide emulsion. The pea protein solution was used without centrifugation.

  The pea protein and soy polysaccharide solutions were each adjusted to pH 7.0, then mixed and stirred for 2 hours. The protein concentration was 5 mg / mL and the WR was changed from 1: 1 to 1: 6. The mixture was emulsified with a 10% oil fraction at 800 bar at pH 3.5 for 4 minutes, followed by heat treatment at 90 ° C. for 1 hour. The data in Table 26 confirms that an appropriate WR value is in the range of 1: 2 to 1: 6.

Claims (12)

a) 0.1 to 70% by weight of one or more fat-soluble active ingredients based on the composition;
b) one or more plant proteins selected from the group of proteins suitable for food use; and c) a composition comprising one or more soy soluble polysaccharides,
The total amount of the protein and the amount of polysaccharide corresponds to 10 to 85% by weight based on the composition in the dry substance, and the weight ratio of protein to polysaccharide is selected to be 1: b. Wherein b comprises 0.5-15.   The fat-soluble active ingredient (s) are vitamins A, D, E, K, and derivatives thereof; polyunsaturated fatty acids; lipophilic health ingredients; carotenoids; and flavorings or fragrances and their 2. Composition according to claim 1, characterized in that it is selected from the group consisting of mixtures.   The fat-soluble active ingredient (s) is a carotenoid, in particular β-carotene, lycopene, lutein, bixin, astaxanthin, apocarotenal, β-apo8′-carotenal, β-apo-12′-carotenal, canta The composition according to claim 1, wherein the composition is xanthine, cryptoxanthin, citranaxanthin and / or zeaxanthin.   Said lipophilic health ingredient (s) is resveratrol; senkyu; ubiquinones and / or ubiquinol (s), preferred coenzyme Q10, coenzyme Q9, and / or Composition according to claim 2, characterized in that they are selected from the group consisting of their reduced form (corresponding ubiquinol); genistein and α-lipoic acid.   5. The protein according to claim 1, wherein the protein is a plant protein (one or more compounds) selected from the group consisting of soybean protein, lupine protein, pea protein, and potato protein. The composition according to one item.   6. Composition according to claim 5, characterized in that the plant protein is soybean or pea protein. I) suspending the protein in water;
II) optionally removing undissolved protein from the suspension of step I);
III) mixing one or more soy soluble polysaccharides in a weight ratio of 1: 0.5 to 1:15 protein and polysaccharide;
IV) adjusting the pH to a value comprised between 3 and 5;
V) adding an organic phase comprising one or more lipophilic active ingredients to the composite;
VI) homogenizing the mixture of step V) by conventional emulsification processes known to those skilled in the art;
VII) heating the emulsion at a temperature comprised between 70 and 95 ° C, preferably between 80 and 90 ° C for at least 45 minutes, preferably at least 1 hour;
VIII) optionally, drying the emulsion of step VII).   A plant protein-soybean soluble polysaccharide complex obtained by the method according to claim 7.   The plant protein-soybean soluble polysaccharide according to claim 8, wherein the plant protein is soybean protein or pea protein.   Use of the composition according to any one of claims 1 to 6 for enrichment, enhancement and / or coloring of food, beverage, animal feed, cosmetic or pharmaceutical composition.   A method for producing a beverage, comprising the step of mixing the composition according to any one of claims 1 to 6 with further usual ingredients of the beverage.   A beverage obtainable by the method according to claim 11.